2. 218 Christoph Spötl
2. Description of the method
2.1 Preparation steps
The method employs 10 ml septum-capped Na-glass vials (Labco exetainer) that can be anal-
ysed using an autosampler. Prior to going into the field, the exetainer is preloaded with five
droplets of phosphoric acid (ca. 90%), capped and the headspace is flushed with He 6.0 in the
autosampler by penetrating the butyl rubber septa of the exetainers’ disposable caps (Labco
catalogue no. VC301) using a double-hole needle. Alternatively, the acid can also be injected
at the sampling site (by penetrating the septum using a needle syringe) prior to introducing
the water sample. At the sampling site, an aliquot of the water sample is injected into the
exetainer using a needle syringe. Care should be taken to avoid small air bubbles that become
trapped in the syringe while the sample is drawn up. The amount of sample injected depends
on the carbonate alkalinity and the dynamic range of the mass spectrometer. Table 1 gives an
overview of the volumes used for samples that have been analysed in our laboratory at the
University of Innsbruck. The acid-water reaction begins immediately upon injection. Ideally,
the exetainer should remain in upright position afterwards in order to avoid droplets of solution
on the bottom of the rubber septa, which can give rise to water being entrained into the mea-
surement system. In our experience, however, less careful handling of samples (e.g. shipping
by courier) does not pose a problem, if the septa are carefully checked using a lamp prior to
the measurement (occasional droplets can be effectively removed by gently knocking against
2.2 Isotopic analysis
The CO2 in the headspace of the exetainer is analysed within a few days after injection. In
the laboratory, the exetainers are loaded into the CTC Combi-Pal autosampler linked to a
Gasbench II. Alternatively, exetainers can be analysed manually by putting one by one on
the measurement needle of the Gasbench. Once penetrated by the double-hole needle, sample
gas (He plus CO2 and small amounts of N2 degassed from the sample) is pushed out of
the exetainer by a slow flow of He 6.0 from the Gasbench. The gas is subsequently purified
(Nafion trap to remove water vapour) and separated gas chromatographically using a Poraplot
Q GC column (fused silica tubing, 25 m × 0.32 mm, kept at 70 ◦
C), followed by a second
Nafion trap and then leaked into the source of the Deltaplus
XL mass spectrometer. The entire
procedure is fully automated and implemented into the measurement protocol under ISODAT.
Each measurement commences with three injections of CO2 reference gas, followed by nine
injections of sample gas via an 8-portValco valve. The software calculates the raw δ13
Table 1. Approximate sample volumes required for DIC analysis at
the University of Innsbruck Stable Isotope Laboratory, which utilizes a
DeltaplusXL mass spectrometer (dynamic range 0–10V m/z 44).
Carbonate hardness (◦dH)† HCO3 (mg/l) Sample volume (µl)
1–2 22–44 1500
2–5 44–109 700
5–8 109–174 500
8–11 174–240 300
11–15 240–327 200
15–20 327–436 100
Quoted in degrees German hardness.
3. 13CDIC in ground waters 219
by taking the average of the nine consecutive sample peaks and comparing their raw ratios to
those of the reference gas.
2.3 Calibration against VPDB
Because there are no durable DIC standards available for stable isotope analysis, we use the
following procedure to calibrate the raw delta values against the VPDB: DIC samples are
run intermittently with calcite samples (DIC is measured during the working day and calcite
runs overnight). We use the calcite standard δ13
C values of the runs before and after a DIC
measurement to correct and calibrate the δ13
CDIC values (that can be traced back to NBS
and IAEA carbonate standards). A linearity correction is performed using the slope δ13
intensity of m/z 44 as measured during the calcite runs . Alternatively, the calibration can
be performed by injecting CO2 of known isotopic composition (e.g. ISO TOP CO2 of Messer)
into exetainer vials and by performing a linearity correction and calibration based on these
3. Results and discussion
Internal precision of δ13
C is typically between 0.05 and 0.08 ‰, depending on the CO2 content
of the sample and the linearity (m/z 44 vs. raw δ13
C) of the system. External precision was
measured twice within a period of 2 years by preparing nine and 20 aliquots of laboratory tap
water, respectively. The standard deviation was <0.08 ‰ in both test runs (table 2). Previous
tests with our instrument (using carbonate samples) have shown that precision deteriorates
rapidly if the intensity of the first sample peak (m/z 44) falls below ∼0.5V. It is therefore
important to adjust the amount of sample injected into the exetainer (cf. table 1).
The accuracy of our system was explored by preparing artificial DIC samples of Na2CO3
(Merck) whose C isotopic composition was measured using the classical reaction with dry
orthophosphoric acid. Dry soda powders may experience changes in their δ13
C value during
long storage, if they are in continuous contact with CO2 [9, 10]. In addition, alkaline solutions
absorb CO2 giving rise to kinetic C isotope fractionation . We minimized these effects by
preparing Na2CO3 DIC solutions and Na2CO3 samples for the H3PO4 reaction rapidly from the
same, homogenized aliquot of solid Na2CO3. Five DIC samples were prepared by dissolving
∼40 mg in 100 ml of freshly produced deionized water (electric conductivity 0.08 µS/cm).
Dissolution was accomplished within minutes and 1 ml each of these solutions was injected
into two He-flushed exetainer each preloaded with five droplets of H3PO4. The entire process
was completed in <1 h, and the DIC samples were analysed after 1 h.At the same time, aliquots
of the same Na2CO3 powder were deposited in 10 ml borosilicate glass vials (between 300
and 400 µg each), capped and flushed with He within <2 h. The latter batch of samples was
reacted with H3PO4 at 72 ◦
C together with a set of in-house calcite standards whose isotopic
composition can be traced back to NBS and IAEA standard reference materials . The
solid Na2CO3 samples yielded a mean δ13
C value of −4.75 ± 0.10 ‰ (n = 12). This value
is only marginally higher than the mean δ13
CDIC value: −4.92 ± 0.08 ‰ (n = 10). These two
data groups are indistinguishable within their 1 σ analytical error margins. The fact that the
CDIC value is slightly lower than the C isotopic composition of the solid product actually
4. 220 Christoph Spötl
Table 2. Tests of precision using laboratory tap water.
Test 1 Test 2
Exetainer no. m/z 44 δ13C (‰) Exetainer no. m/z 44 δ13C (‰)
1 4.03 −3.90 1 5.77 −4.06
2 3.97 −3.89 2 5.87 −4.07
3 4.15 −3.79 3 5.59 −4.08
4 4.07 −3.78 4 5.82 −4.07
5 4.04 −3.77 5 5.45 −4.15
6 4.10 −3.69 6 5.96 −3.98
7 4.10 −3.74 7 5.78 −4.02
8 4.01 −3.85 8 6.49 −3.93
9 4.14 −3.79 9 4.08 −3.99
10 4.03 −3.78
11 4.05 −3.84
12 4.09 −3.79
13 4.13 −3.84
14 4.08 −3.86
15 4.00 −3.86
16 3.96 −3.87
17 4.01 −3.86
18 4.05 −3.85
19 4.08 −3.82
20 4.02 −3.82
Mean −3.82 Mean −4.04
SD 0.05 SD 0.07
Test 2 was performed ∼2 years after test 1. The intensity of mass 44 of the first sample peak is
air during solution preparation; otherwise its δ13
C value would have increased (the δ13
of the laboratory air value is ca. −10 ‰ and would give rise to a δ13
CDIC value of ∼9 ‰
higher than that in case of complete exchange).
3.3 Storage effects
Previous studies using glass autosampler vials with butyl rubber caps reported evidence of
Oand – toalesserextentalso – intheδ13
the CO2 content and its isotopic composition [13–16]. We have systematically studied storage
effects of Labco autosampler vials and found only insignificant (i.e. <0.1 ‰) C isotope shifts
within a period of up to 2 weeks of storage at room temperature . We routinely ship He-
flushed exetainer to foreign field locations by courier, where phosphoric acid is being injected
followed by injection of an aliquot of the water sample. The exetainer is shipped back to the
laboratory (again by courier) and analysed within 2 days. Blank exetainer vials (with and
without acid) are routinely included in these sample batches, but no evidence of significant
leakage was found. We therefore conclude that the butyl rubber septa are gastight, provided
that the caps are properly tightened. In addition, the relative insensitivity of the C isotopic
composition of DIC samples in exetainer to minor gas exchange over periods of up to 2 weeks
reflects the much higher pCO2 in the exetainer when compared with the outside air pCO2.
Nevertheless, we recommend to keep the time interval between flushing the samples, injection
the water and analysing them as short as possible (i.e. within 1 week) and to include blank
exetainer within each sample batch as monitors of possible gas leakage. The use of Kel-F discs
in addition to butyl rubber septa [cf. ref. 16] is not applicable for the technique presented in this
paper, because Kel-F discs loose their sealing function once they are penetrated by a needle.
5. 13CDIC in ground waters 221
Given that the total analysis time is 10 min per sample, the method described earlier is
well suited for large sample batches run by an autosampler. Its advantages include minimal
sample handling in the field, avoidance of toxic substances, small sample volumes, automated
continuous-flow online sample preparation and analysis using an autosampler. The method
has so far been used primarily to analyse water samples from karst aquifers. Its application to
other water types (e.g. very low-DIC or high-DOC surface water samples) appears feasible,
but should be thoroughly tested first.
This work has been supported by the Austrian Science Funds under grant STARTY122-GEO.
I would like to thank my lab manager, Manuela Wimmer, and Anna F. Tooth, Karl-Heinz
Offenbecher, Ronny Boch and Michael Meyer for their support in the field and the laboratory
and two referees for their helpful comments.
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